The mutation rate of turnip mosaic virus (TuMV) was estimated using the Luria-Delbrück fluctuation test (1). One hundred susceptible Arabidopsis thaliana plants were inoculated with sap containing TuMV and maintained in a growth chamber until the virus colonized the plants systemically. From each of these plants, virus was extracted from symptomatic tissue 14 days post-inoculation. The fact that only a fraction of the virus-infected host tissues was extracted was taken into account when calculating the mutation rate. Extracted virions were challenged against the TuMV-resistant A. thaliana line 12-4 expressing an artificial microRNA (amiRNA). From the 100 wild- type A. thaliana plants, 11 contained escape mutants that were capable of infecting 12-4 plants. A maximum-likelihood algorithm was used to estimate of the mutation rate for the amiR target locus, yielding m = 5.55 × 10⁻⁵ s/r. Assuming that resistance breaking can be conferred by every single substitution in the 21 nucleotide amiRNA target and assuming that 40% such substitutions are lethal to the virus, T_s = 21 ´ 3 ´ 0.4 = 25.2. Therefore, m_s/n/r = 5.55 × 10⁻⁵ ´ 3 / 25.2 = 6.6 ´ 10^-6.

1. de la Iglesia, F., F. Martínez, J. Hillung, J.M. Cuevas JM, P.J. Gerrish, J.A. Daròs, and S.F. Elena. 2012. Luria-Delbruck estimation of turnip mosaic virus mutation rate in vivo. J. Virol., in press.